Tissue phenotype depends on reciprocal interactions between the extracellular matrix and the structural organization of the nucleus

What determines the nuclear organization within a cell and whether this organization itself can impose cellular function within a tissue remains unknown. To explore the relationship between nuclear organization and tissue architecture and function, we used a model of human mammary epithelial cell acinar morphogenesis. When cultured within a reconstituted basement membrane (rBM), HMT-3522 cells form polarized and growth-arrested tissue-like acini with a central lumen and deposit an endogenous BM. We show that rBM-induced morphogenesis is accompanied by relocalization of the nuclear matrix proteins NuMA, splicing factor SRm160, and cell cycle regulator Rb. These proteins had distinct distribution patterns specific for proliferation, growth arrest, and acini formation, whereas the distribution of the nuclear lamina protein, lamin B, remained unchanged. NuMA relocalized to foci, which coalesced into larger assemblies as morphogenesis progressed. Perturbation of histone acetylation in the acini by trichostatin A treatment altered chromatin structure, disrupted NuMA foci, and induced cell proliferation. Moreover, treatment of transiently permeabilized acini with a NuMA antibody led to the disruption of NuMA foci, alteration of histone acetylation, activation of metalloproteases, and breakdown of the endogenous BM. These results experimentally demonstrate a dynamic interaction between the extracellular matrix, nuclear organization, and tissue phenotype. They further show that rather than passively reflecting changes in gene expression, nuclear organization itself can modulate the cellular and tissue phenotype.

Reciprocal interactions between β1-integrin and epidermal growth factor receptor in three-dimensional basement membrane breast cultures: A different perspective in epithelial biology

Anchorage and growth factor independence are cardinal features of the transformed phenotype. Although it is logical that the two pathways must be coregulated in normal tissues to maintain homeostasis, this has not been demonstrated directly. We showed previously that down-modulation of β1-integrin signaling reverted the malignant behavior of a human breast tumor cell line (T4–2) derived from phenotypically normal cells (HMT-3522) and led to growth arrest in a three-dimensional (3D) basement membrane assay in which the cells formed tissue-like acini (14). Here, we show that there is a bidirectional cross-modulation of β1-integrin and epidermal growth factor receptor (EGFR) signaling via the mitogen-activated protein kinase (MAPK) pathway. The reciprocal modulation does not occur in monolayer (2D) cultures. Antibody-mediated inhibition of either of these receptors in the tumor cells, or inhibition of MAPK kinase, induced a concomitant down-regulation of both receptors, followed by growth-arrest and restoration of normal breast tissue morphogenesis. Cross-modulation and tissue morphogenesis were associated with attenuation of EGF-induced transient MAPK activation. To specifically test EGFR and β1-integrin interdependency, EGFR was overexpressed in nonmalignant cells, leading to disruption of morphogenesis and a compensatory up-regulation of β1-integrin expression, again only in 3D. Our results indicate that when breast cells are spatially organized as a result of contact with basement membrane, the signaling pathways become coupled and bidirectional. They further explain why breast cells fail to differentiate in monolayer cultures in which these events are mostly uncoupled. Moreover, in a subset of tumor cells in which these pathways are misregulated but functional, the cells could be “normalized” by manipulating either pathway.

Modulation of cholecystokinin activity by albumin.

We found that a variety of cholecystokinin (CCK) receptor ligands bind to bovine serum albumin (BSA). This binding was rapid, fully reversible, temperature independent, of low affinity, and specific for BSA; it depended on the concentration of BSA, the chemical structure of the ligand, and the chemical composition of the incubation medium. BSA also decreased the binding of 125I-labeled CCK octapeptide (125I-CCK-8) to CCK receptors on pancreatic acini and membranes but increased the potency with which CCK-8 inhibited binding of 125I-CCK-8. These counterintuitive findings appeared to result from BSA altering the affinities of CCK-8 for different affinity states of the pancreatic CCK receptor. An alternate hypothesis is that BSA increased the efficacy of CCK-8 such that it bound to receptors and also caused biochemical changes in other receptors that reduced their ability to bind 125I-CCK-8. BSA enhanced the ability of CCK-8 to stimulate amylase secretion from pancreatic acini and to cause contraction of dispersed gastric smooth muscle cells. Thus, CCK can bind to BSA, and the BSA-CCK complex has substantially different activities from the free, uncomplexed hormone.

Method for the generation and cultivation of functional three-dimensional mammary constructs without exogenous extracellular matrix

During puberty, pregnancy, lactation and postlactation, breast tissue undergoes extensive remodelling and the disruption of these events can lead to cancer. In vitro studies of mammary tissue and its malignant transformation regularly employ mammary epithelial cells cultivated on matrigel or floating collagen rafts. In these cultures, mammary epithelial cells assemble into three-dimensional structures resembling in vivo acini. We present a novel technique for generating functional mammary constructs without the use of matrix substitutes.

Desarrollo de la ovotestis del "caracol africano" achatina fulica (achatinidae) en el Valle del Cauca [recurso electr?nico]

RESUMEN : En Colombia se ha reportado en 111 municipios de 24 departamentos la presencia del caracol africano Achatina fulica, clasificada como una de las 100 especies m?s da?inas del mundo. Debido a que es plaga de diversos cultivos agr?colas y potencialmente peligrosos para la salud humana, ha sido estudiado en varios aspectos de su biolog?a, entre ellos la reproducci?n. Sin embargo, ?ste es el primer trabajo en el cual se caracterizan las etapas de desarrollo de la ovotestis y su maduraci?n, con base en una poblaci?n de 60 individuos de cuatro municipios del Valle del Cauca. Los muestreos se realizaron entre los meses de octubre y diciembre del 2013. Para ?sta caracterizaci?n se emplearon t?cnicas histol?gicas usando hematoxilina-eosina. Se describieron las diferentes estructuras presentes, desde la formaci?n de los primeros acinos de ovotestis hasta el vaciado de espermatozoides y ovocitos, se cuantificaron y midieron los acinos y los ovocitos. Estos ?ltimos, se caracterizaron a trav?s de la morfolog?a, el tama?o y la ubicaci?n en el acino, clasific?ndolos como ovocitos premei?ticos, previtelog?nicos y vitelog?nicos. Con la informaci?n obtenida y los estados de la espermatog?nesis, se definieron cinco etapas de desarrollo: inmaduros, inicio de gametog?nesis, madurez temprana, madurez total y vaciado, siendo excluyentes entre s?. Las poblaciones estudiadas se compararon mediante la cantidad de los ovocitos en los diferentes estados de madurez y el ?ndice de madurez ovoc?tica, sin diferencias significativas existentes, lo cual sugiere que presentan el mismo patr?n de reproducci?n para esta ?poca del a?o. ABSTRACT : Achatina fulica, which is considered one of the top hundred most dangerous species on the world, has been reported in 111 municipalities in 24 departments of Colombia. Because this species is a plague to several crops and potentially harmful to human health, several aspects of its biology have been studied, including it?s the reproductive biology. However, this is the first work where the ovotestis developmental stages and maturation are characterized. This was accomplished using 60 specimens from four municipalities of Valle del Cauca department. Samplings were conducted between monts october and december 2013. For the characterization, histological techniques using hematoxilin-eosin were used. The different structures were carefully described, from the formation of the first ovotestes? acini to the sperm emptying; the acini and ovocites were counted and measured. The latter were characterized, using morphology, size and the position on the acini, as pre-meiotic, pre-vitellogenic and vitellogenic oocytes. Based on this information and the spermatogenesis stages, five developmental stages were defined: immature, early gametogenesis, early maturity, mature and emptiness, which were mutually exclusive. The studied populations were compared using the amount of oocytes in the different maturity stages and the oocytic maturity index, but no significant differences were found, which suggests that the all the populations show the same reproductive pattern during this year season.

Gross and microscopic observations on the lingual structure of the franciscana (Pontoporia blainvillei-gervais and d'orbigny, 1844)

In most anatomical studies developed with mammals, the tongue is described as highly differentiated among different species. However, studies on the tongue of aquatic mammals are still limited as compared to those on terrestrial mammals. The aim of this study was to describe the tongue morphology of the Franciscana dolphin (Pontoporia blainvillei) using macroscopic observations, light, and scanning electron microscopy. Microscopically, the dorsal surface was covered by a keratinized stratified epithelium. Salivary gland acini were found on the middle and caudal third of the tongue. The dorsal surface was totally covered by filiform papillae with a connective tissue core and a connective tissue structure round in shape in the middle and caudal regions. Microsc. Res. Tech. 75:737742, 2012. (C) 2012 Wiley Periodicals, Inc.

Distribution and expression of leptin in major salivary glands of a hyperleptinemia animal model

Saliva production is mainly regulated by the autonomic nervous system (sympathetic and parasympathetic); however studies indicate a possible hormonal influence on the control of salivary secretion. This study aims to assess if the induction of increased levels of circulating leptin influence the immunohistochemical expression of leptin at the level of major salivary glands in Wistar rats. It was found that the expression, in qualitative terms, of leptin has been positive, being more evident in submandibular and sublingual glands, either in the acini or ducts. However, through this technique, no obvious differences between groups could be observed. The results suggest that circulating leptin levels may not affect the expression of this hormone in the major salivary glands.